Analogs of the tripeptide hormone TRH, in which H-2 or H-4 of the imidazole ring has been replaced by a trifluoromethyl group, show pituitary and cardiovascular activities equal to, or better than, those of the natural peptide in the conscious rat. Thus, the loss of imidazole basicity and the introduction of a bulky substituent at either position seems to be unimportant in ligand recognition, Yet, the imidazole ring cannot be irrelevant to recognition since the 4-nitro analogue shows no pituitary activity but retains full cardiovascular activity. We have demonstrated the duality (or multiplicity) of TRH receptors in brain and pituitary. The further demonstration of separation of activities by our novel analogues offers an opportunity to attempt clinical removal of the set of activities governed by one receptor or another. (Trifluoromethyl)imidazoles show the very unique property of losing HF at mildly alkaline pH to form extremely electrophilic difluorodiazafulvenes. The generation of the latter species within a receptor's ligand-binding site may result in irreversible bond formation with a protein nucleophile in the binding site. At 37 degreesC and pH 8.5, the half-time for fulvene generation from 2-CF3-Im-TRH is ca. 30 hr. It is unreasonable, therefore, to search for irreversible binding to pituitary or brain tissue during the period of a normal incubation. Analysis of pK and reactivity data for a large number of additionally substituted (trifluoromethyl)imidazoles provided the basis for predicting appropriate extra substituents to give TRH analogues with half-lives ranging from 4 sec to 1.5 yr. We have chosen acetamido as the ideal substituent for our purposes and are now engaged in development of synthetic routes to 4-acetamido-2-trifluoromethyl-Im-TRH. Should this compound show the expected reactivity in binding irreversible to specific TRH receptors in tissue homogenates, it will then be administered to rats and followed by a dose of normal TRH. Strong diminution of either pituitary or CVS activity is expected. Similar approaches are being used to create selective irreversible inhibitors of histamine H1 and H2 receptors.